This invention relates to a method of rapid and precise analysis, both quantitatively and qualitatively, for carbohydrates, and in particular, this invention relates to a method for regular syrups, fructose syrups, and blends of syrups or carbohydrates with sucrose to determine constitutional makeup.
There have been four main procedures in the past for the quantitative analysis of fructose and dextrose in syrups or for the analysis of other syrup and starch compositions.
(1) The method of polarimetry is described in U.S. Pat. No. 3,694,158 and is believed to be the first feasible system for the continuous analysis of a process stream to detect the amounts of dextrose and fructose in mixtures simultaneously. This procedure has the disadvantage, however, of producing imprecise results if the composition contains appreciable amounts (over two percent) of higher homologs of dextrose, i.e. degree of polymerization of 2-10 or higher, e.g. maltooligosaccharides. Furthermore, the method becomes inaccurate if the tested composition contains mixtures of three or more types of sugars. The method is also sensitive to variations in temperature, thus requiring a controlled atmosphere.
(2) High pressure liquid chromatography has been developed to where it is capable of analyzing carbohydrate solutions containing fructose, dextrose, higher homologs of these sugars, and still other sugars. The method is capable of providing an automatic analysis in a period of time of 10-30 minutes. A discussion of this procedure may be found in AACC Paper No. 48, Oct. 5-8, 1976, Annual Meeting In New Orleans, La. by H. D. Scobell. Other than the inordinate length of time to obtain the analysis results, this process appears to be a suitable technique except where there are blends containing sucrose or maltose, or where there are appreciable quantities of maltulose, etc. These components elute simultaneously with other constituents of the analyzed sample and yield imprecise results.
(3) Gas/liquid chromatography is an earlier procedure than high pressure liquid chromatography and it is capable of quantitative analysis of a carbohydrate composition to determine the various sugars present in one analytical scan. The difficulty with this procedure is that it requires a precise technique in the preparation of samples and in the derivatization of samples. Furthermore, the analytical time, not including the sample preparation time, is inordinantly long, i.e. about 20 minutes. See K. M. Brobst and C. E. Lott, Am. Soc. Brew. Chem. Proc. 71-75 (1966).
(4) There are many chemical procedures for the quantitative analysis of dextrose, fructose, higher sugars, and mixtures of sugars. Illustrative of such analytical techniques are:
(a) Cystein carbazole method for the determination of ketose sugars, Corn Refiners Tentative Method E-1, Jan. 6, 1976; PA1 (b) "The Quantitative Determination of Glucose, Fructose, and Sucrose in Fruits and Potatoes", E. S. Della Monica et al, J. Food Sci. 39, pp. 1062-3 (1974); PA1 (c) "Spectrophotometric Analyses Of Glucose And Mixture Of Glucose, Fructose, And Sucrose", E. Garrett and J. Young, J. Pharm, Sci. 58, pp. 1224-7 (1969); and PA1 (d) "Dextrose Equivalent", PA1 (2) Cell Thickness -- 12.5, 15, or 25 microns PA1 (3) Number Of Scans -- 250, 125, 30, and 1 PA1 (4) Resolution -- 4 wave members PA1 (5) Word No. -- 32 bit words PA1 (6) Instrument -- Digilab IRFTS 15 equipped with a double beam head; Perkin Elmer 283 with 580 infrared Spectrophotometer; or Nicolet 7199 FT-FR System PA1 (7) Light -- monochromatic, e.g. by employing filters, lasers, etc. PA1 (8) Concentration -- 10-30% solids preferred, wider ranges if desired PA1 (9) Sample Preparation -- Anomeric equilibrium attained prior to analysis of sample. Equilibrium obtained either by allowing solution to reach equilibrium by natural action or by the addition of several drops of ammonium hydroxide when speed is necessary. PA1 (10) Analysis Of Unknowns -- Samples of fructose corn syrup and regular corn syrup analyzed by high pressure liquid chromatography for carbohydrate composition and by CRA Method E-26 for DE in regular corn syrups.
Corn Refiners Method E-26. These are all wet chemical analyses which have substantially the same disadvantages of the possibility of technician error, the requirement of large amounts of bench space and analytical equipment, and the large amount of time involved in the analysis. Furthermore, these techniques are either so specific that they are capable of analyzing only a single type of sugar or they are so non-specific that they are incapable of distinguishing one type of sugar from another.
The present invention provides an analytical process which overcomes these disadvantages. It is extremely fast in that it can provide a complete analysis in 1-3 minutes, frequently less than 1 minute. There is no necessity for derivatization nor is there a need to carefully control the amount of solids in the sample. It is preferable to have anomeric equilibrium in the mixtures analyzed. This technique is capable of analyzing samples containing from less than 1 percent up to 100 percent solids and it is capable of installation on a production line for continuous or semi-continuous analysis of process streams. The technique is capable of handling higher sugars as well as mixtures of sugars. It is capable of providing information as to carbohydrate complex formations, such as fructose-H.sub.3 BO.sub.3. It is also capable of providing an analysis of the degree of anomerization and the rate of anomerization. The process is also able to provide information as to the extent of derivatization and other behavorial questions such as carbohydrate anomerization in acids, bases, and organic solvents.
The present invention is based on infrared (IR) analysis of carbohydrates. While IR spectra of carbohydrates have been studied and characterized for many years [two reviews have been published: "Infrared Spectra of Carbohydrates" by W. Brock Neely, in ADVANCES IN CARBOHYDRATE CHEMISTRY, 12 13-33 (1957) and "Infrared Spectroscopy and Carbohydrate Chemistry" by H. Spedding in ADVANCES IN CARBOHYDRATE CHEMISTRY, 19 23-49 (1964)] it was not immediately apparent that IR could be used as a qualitative/quantitative analytical tool. This position was taken in a paper by W. J. Hoover, et al. in J. FOOD SCIENCE, 30 253-261 (1965) "Isolation and Evaluation of the Saccharide Components to Starch Hydrolysates II. Evaluation" where it was stated with reference to hydrolyzed starch, "No trends were noted in shifts of peaks or development of peaks with increase in molecular weight (from dextrose) . . . The spectra of all of the saccharides were so similar that they could not be used in identifying or distinguishing between the various malto-oligosaccharides." Furthermore, a paper "Infrared Spectra of Carbohydrates in Water and a New Measure of Mutarotation" by Frank S. Parker in BIOCHIM. BIOPHYS. Acta. 42 513-519 (1960) indicates that the absorbance at 1143 cm.sup.-1 is the most indicative band for observing mutarotation of .alpha.- and .beta.-glucose. In accordance with the present invention absorbances at 1038 cm.sup.-1 and 1080 cm.sup.-1 provide much more useful information.
Very little infrared work has been done specifically for fructose. Typical of that work is that which is described in "Identification of the Anhydrides of D-Fructose from the `Fingerprint` Region of their Infrared Spectra" by W. W. Binkley, et. al., INTERNATIONAL SUGAR JOURNAL 73 259-261 (1971), which clearly is not directed at the analytical purposes of the present invention.